Antibody-mediated Gene Therapy for the Treatment of Cancer

抗体介导的癌症基因疗法

• 批准号: 8319671
• 负责人: Tracy Ruth Daniels-Wells
• 金额: $ 14.27万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-28 至 2014-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8319671
• 关键词: 5 fluorouridine; Adverse effects; Affect; Animal Model; Antibodies; Antibody-Directed Enzyme Prodrug Therapy; Applications Grants; Avidin; Award; B lymphoid malignancy; B-Cell Lymphomas; B-Cell NonHodgkins Lymphoma; B-Lymphocytes; Binding; Bystander Effect; Cancer Patient; Cell Line; Cells; Cellular biology; Chimeric Proteins; Cytosine; DNA; Development; Dimensions; Disease; Drug Kinetics; Environment; Enzymes; Evaluation; Faculty; Flucytosine; Fluorouracil; Future; Gene Delivery; Gene Expression; Genes; Goals; Growth; Health; Hematopoietic; Hematopoietic stem cells; Hodgkin Disease; Human; IgG3; Immunoglobulins; Immunotherapy; In Vitro; Institution; Knowledge; Lentivirus Vector; Lymphoma; Magic; Malignant - descriptor; Malignant Neoplasms; Mantle Cell Lymphoma; Mediating; Mentors; Mus; N glycosidase; Non-Hodgkin' s Lymphoma; Normal Cell; Oncogenes; Pain; Pharmaceutical Preparations; Plants; Positioning Attribute; Preparation; Prodrugs; Property; Protein Biosynthesis; Proteins; Rattus; Receptors, Antigen, B-Cell; Reporter Genes; Research; Research Personnel; Role; SCID Beige Mouse; Saponaria; Species Specificity; Sprague-Dawley Rats; Stromal Cells; Surface; Survival Rate; System; Testing; Therapeutic; Toxic effect; Toxin; Training; Transferrin Receptor; Transgenes; Tumor Antigens; Ubiquitin; United States; Uracil phosphoribosyltransferase; Xenograft procedure; Yeasts; base; cancer cell; cancer therapy; career development; cellular pathology; chimeric antibody; cytotoxicity; economic cost; effective therapy; experience; gene therapy; improved; in vivo; novel therapeutic intervention; novel therapeutics; outcome forecast; overexpression; promoter; receptor; receptor mediated endocytosis; selective expression; skills; standard of care; success; targeted delivery; therapeutic gene; transgene expression; tumor; vector

DESCRIPTION (provided by applicant): The proposed studies focus on the development and characterization of new highly targeted gene therapy approaches for the systemic treatment of aggressive B-cell lymphomas with an emphasis on mantle cell lymphoma (MCL). Lymphomas are subdivided into Hodgkin's lymphoma (HL) and non-Hodgkin's lymphoma (NHL), of which in the United States more than 85% are NHL. MCL is an aggressive form of B-cell NHL with a very poor prognosis. MCL comprises 5-10% of NHL cases, has a median survival of about 4 years, and a long- term survival rate of less than 15%, which has not significantly changed in the past 20 years. Currently, there is no accepted standard of care for the treatment of MCL and the disease is considered incurable. Therefore, novel therapeutic approaches are urgently needed. In the strategies described in this application, MCL targeting will occur via two mechanisms: 1) through the targeting of a tumor-associated antigen (TAA) on the surface of malignant B cells and 2) through the selective expression of toxic genes using a cell-specific promoter. TAA on the surface of cancer cells serve as excellent targets for immunotherapy. Therefore, the first level of my targeted strategy will occur through the use of a mouse/human chimeric antibody-avidin fusion protein specific for the transferrin receptor (TfR). This receptor is an attractive target for cancer therapy due to its elevated expression on the surface of cancer cells, its ability to internalize, and its central role in the cellular pathology of cancer. However, the TfR is expressed on some normal cells at various levels. In order to further improve malignant cell targeting, the second level of my targeted strategy focuses on limiting the expression of toxic genes to malignant cells by using the immunoglobulin promoter. The central hypothesis of the present proposal is that TfR overexpression on the surface of MCL can be used as an effective target for TfR- mediated gene delivery, for which the transgene will be transcriptionally restricted. Since tumor targeting will occur on two levels, I also hypothesize that this strategy will be extremely effective in eliminating malignant B cells in vivo without the severe side effects that limit the efficacy of most cancer therapeutics. The antibody-avidin fusion protein that targets the TfR is a unique drug since it serves as a universal delivery system for a wide variety of biotinylated agents. The antibody-avidin fusion protein will be conjugated to either biotinylated DNA or biotinylated lentiviral vectors in order to deliver a toxic gene into malignant B cells by receptor-mediated endocytosis. The use of two independent and non-exclusive gene therapy strategies is proposed in this application. The first gene encodes the toxin saporin, a ribosomal inactivating protein that is derived from the plant Saponaria officinialis. Saporin is a single chain toxin that cannot enter cells by itself due to the lack of a cell-binding domain. Saporin is a highly toxic and once inside the cell it inhibits protein synthesis through its N-glycosidase activity that leads to the inactivation of the 28S ribosomal subunit. The second gene that will be used encodes a chimeric yeast enzyme (FCU1) that consists of cytosine deaminse (CD) and uracil phosphoribosyltransferase (UPRT). This enzyme converts the prodrug 5-fluorocytosine to the toxic metabolites 5-fluorouracil (5-FU) and 5-fluorouridine 5'monophosphate (5-FUMP) and thus is an antibody- directed enzyme prodrug therapy (ADEPT) approach. The prodrug will be converted to its toxic metabolites within the tumor microenvironment. It is expected there will be a bystander effect associated with ADEPT therapy since the toxic metabolites can be released from targeted cells and taken up by non-targeted malignant cells in the tumor environment as well as stromal cells that support the growth of the malignant cells. Importantly, these two strategies can be used in the future in combination to maximize their anti-tumor effects. The use of this dual targeting strategy using either toxic gene is expected to increase the anti-tumor activity compared to singularly targeted agents, as well as eliminate the potential systemic toxicity of the treatment. To execute this project I propose three specific aims: Aim 1: Reporter gene vector construction and in vitro optimization of gene delivery. Aim 2: Toxic gene vector construction and in vitro evaluation of targeted anti-cancer activity. Aim 3: Evaluation of toxicity, pharmacokinetics, and anti-tumor activity in animal models. This project is expected to result in important advances not only in the fields of cancer gene therapy and treatment of MCL, but also in my career development. In fact, many new skills will be acquired that will increase my knowledge and research experience. This training will aid in the future preparation of grant proposals that will allow me to become a better candidate for an academic faculty position at a leading institution, which is my long-term goal. It is my goal to become an independent investigator to better understand cancer cell biology in order to develop new therapeutics that will help reduce the pain and suffering encountered by cancer patients. The outstanding research environment at UCLA, the guidance from my experienced mentors, and this award will greatly facilitate my success in reaching my goals and will open a new dimension in my professional development.

描述（由申请人提供）：拟议的研究重点是开发和表征新的高度靶向基因治疗方法，用于全身治疗侵袭性b细胞淋巴瘤，重点是套细胞淋巴瘤（MCL）。淋巴瘤又分为霍奇金淋巴瘤（HL）和非霍奇金淋巴瘤（NHL），其中在美国85%以上为NHL。MCL是一种侵袭性b细胞NHL，预后很差。MCL占NHL病例的5-10%，中位生存期约为4年，长期生存率低于15%，近20年来没有明显变化。目前，对于MCL的治疗没有公认的护理标准，这种疾病被认为是无法治愈的。因此，迫切需要新的治疗方法。在本应用中描述的策略中，MCL靶向将通过两种机制发生：1)通过靶向恶性B细胞表面的肿瘤相关抗原（TAA）和2)通过使用细胞特异性启动子选择性表达毒性基因。癌细胞表面的TAA是免疫治疗的良好靶点。因此，我的靶向策略的第一级将通过使用小鼠/人嵌合抗体-亲和素融合蛋白特异性转铁蛋白受体（TfR）来实现。由于其在癌细胞表面的高表达，其内化能力以及其在癌症细胞病理中的核心作用，该受体是癌症治疗的一个有吸引力的靶点。然而，TfR在一些正常细胞上有不同水平的表达。为了进一步提高恶性细胞的靶向性，我的靶向策略的第二个层次是利用免疫球蛋白启动子限制毒性基因对恶性细胞的表达。本研究的核心假设是，MCL表面的TfR过表达可以作为TfR介导的基因传递的有效靶标，因此转基因将受到转录限制。由于肿瘤靶向将在两个层面上发生，我也假设这种策略将在体内非常有效地消除恶性B细胞，而不会产生限制大多数癌症治疗效果的严重副作用。靶向TfR的抗体-亲和素融合蛋白是一种独特的药物，因为它可以作为多种生物素化药物的通用递送系统。